1. Field of the Invention
This invention relates to the detection of small magnetized particles by a magnetic sensor, particularly when such particles are attached to molecules whose presence or absence is to be determined in a chemical or biological assay.
2. Description of the Related Art
Magnetic devices have been proposed as effective sensors to detect the presence of specific chemical and biological molecules when, for example, such molecules are a part of a fluid mixture that includes other molecules whose detection is not necessarily of interest. The basic method underlying such magnetic detection of molecules first requires the attachment of small magnetic (or magnetizable) particles to all the molecules in the mixture that contains the specific molecules to be detected. Because of their small size these attached particles are “super-paramagnetic”, meaning they ordinarily retain no meaningful magnetic moment. However, when placed in an external magnetic field, these particles develop an induced magnetic moment and can produce a corresponding magnetic field, which we will call a “strayfield” herein.
The magnetic particles are made to attach to the molecules in the mixture by coating the particles with a chemical or biological species that binds (e.g. by covalent bonding) to those molecules. Then, a surface (i.e., a substrate) is provided on which there has been affixed receptor sites (e.g. specific molecules) to which only the target molecules (the molecules whose presence is to be measured) will bond. After the mixture has been placed in contact with the surface so that the target molecules have bonded to it, the surface can be flushed in some manner to remove all unbound molecules. Because the bonded target molecules are equipped with the attached magnetic particles, it is only necessary to detect the magnetic particles to be able, at the same time, to assess the number of captured target molecules. Thus, the magnetic particles are simply “flags,” which can be easily detected (and counted) once the target molecules have been captured by chemical bonding to the receptor sites on the surface. The issue, then, is to provide an effective method of detecting the small magnetic particles, since the detection of the particles is tantamount to detection of the target molecules.
One prior art method of detecting small magnetic beads affixed to molecules bonded to receptor sites is to position a magnetic sensor device beneath them; for example, to position it beneath the substrate surface on which the receptor sites have been placed.
FIG. 1 is a highly schematic diagram (typical of the prior art methodology and also exemplary of a portion of the present invention) showing a magnetic (i.e. magnetizable) particle (10) covered with receptor sites (20) that are specific to bonding with a target molecule (30) (shown shaded). The target molecule is shown as bonded to one of a plurality of identical sites (50) affixed to a substrate (70). The substrate (70) is covered with such receptor sites (50) that are also specific to the target molecule (30) and those sites should, in general, be different from the sites that bond the magnetic particle to the molecule. In general, the sites (20) and (50) are capable of bonding to different regions of the target molecule (30). The target molecule (30) is shown bonded to one of the receptor sites (50) on the surface. The magnetic particle (10), being super-paramagnetic because of its small size, must be magnetized by an external field (750) that is directed vertically downward, so that the induced magnetization of the particle (760) (shown as a downward directed arrow) has an external strayfield (95) with a component in the plane of the sensor substrate.
Referring to FIG. 2, there is shown a prior art magnetic sensor (60), similar to a structure used in magnetic random access memory (MRAM), that can be positioned beneath the receptor site of FIG. 1. As shown schematically in the cross-sectional view of FIG. 2, the prior art sensor (60) is based on a magnetic tunneling junction (MTJ) cell, that includes a magnetized “free” layer (61) whose magnetization direction (610) is free to move and a magnetized “pinned” layer (63) whose magnetization (630) is fixed in direction. The two layers are separated by a thin, non-magnetic and electrically non-conducting layer (62), the tunneling barrier layer. The sensor is incorporated within a circuit that can detect changes in the magnetic direction of the free layer relative to the pinned layer, by sensing the changes in the resistance of the sensor, which change is a function of the change in their relative directions. Typically, the MTJ cell is formed so that it has some degree of magnetic anisotropy which provides its magnetization directions (610) with some degree of stability against thermal perturbations and random magnetic fields.
An exemplary circuit includes a selection transistor (70) having a source region (72) to which the sensor element (60) is electrically connected (65), a gate region (74) over which runs a conducting wordline (200) that can effectively activate the gate and allow a sensing current between the source (72) and a grounded (85) drain (76). An electrically conducting bitline (100) contacts the top surface of the sensor to external circuitry and can provide the sensing current that passes between source and drain, thereby effectively measuring the resistance of the sensor.
Referring now to FIG. 3, there is shown an overhead view of the sensor in FIG. 2. By patterning the shape of the sensor into an elliptical form, whereby the sensor has a long dimension and a short dimension, the sensor can operate efficiently as a two-state or “bi-stable” device. The long dimension of the sensor defines an easy axis of magnetization, along which it is energetically favorable the sensor to retain its direction of magnetization. The two labeled directions of magnetization, M1 and M2, are therefore stable, forming the bi-stable state, and serve to store binary information. When the sensor is used as a memory element, as in MRAM devices, these two easy axis directions of magnetization serve as the storage directions because they define positions of stable equilibrium which are unlikely to be disrupted by thermal effects or random magnetic perturbations. Similarly, when such a cell is used as a detector of the presence of magnetized particles (as in an embodiment of the present invention), the two stable equilibrium magnetization directions are stable against perturbations not associated with the presence of a proximate detectable particle. Nevertheless, the cell must be sufficiently sensitive to register the presence of a proximate particle by flipping from one stable equilibrium state to the other.
The short dimension of the sensor defines a hard axis of magnetization, along which the direction of magnetization can be in either of two positions of unstable equilibrium, M4 and M5. These positions will tend to revert to M1 and/or M2 when properly perturbed, as by an external small magnetic field. This lack of stability is a reason why these directions are not used as storage directions. The aspect ratio of the elliptical shape determines Hk, the magnetic anisotropy produced by the shape anisotropy of the ellipse.
Referring to FIG. 4, there is shown an asteroidal curve that defines phase boundaries for the two equilibrium storage states and the non-equilibrium states of such an MTJ cell. The vertical axis measures the magnitude of an external field component, denoted Hy, directed along the cell's hard axis (h.a.). The horizontal axis measures the magnitude of an external field component, denoted Hx, directed along the cell's easy axis (e.a.). M1 and M2 denote the vector magnetizations of the cell in either of the two stable equilibrium positions along the easy axis directions. M3 denotes a generic magnetization vector of the cell when it is directed along a direction θ as the result of the effect of an arbitrary external field having the components (Hx, Hy). The magnitude of θ is given by:sin θ=Hy/Hk where Hk is the shape anisotropy. If Hy=Hk, the magnetization vector will be aligned with the hard axis. On turning off the external field completely, the state of magnetization will be unstable and even the slightest perturbing field will cause it to revert to a stable equilibrium direction along the easy axis. An opposing field along the easy axis direction will cause the magnetization to reverse direction to the other easy axis equilibrium state. This irreversible process will happen when the field component along the easy action direction, Hx, is at the switching threshold, Hc, which has the magnitude Hc=Hk. This relationship assumes that the external magnetization is rotating uniformly past the hard axis energy barrier. In practice this does not happen and when the rotation of the external magnetic field is non-uniform, the switching barrier is actually lower than Hk and is found, experimentally, to be approximately Hk/2. This value determines the stability of stored information.
In a biosensing (magnetic particle sensing) environment, the same sensor serves to detect the presence or absence of a proximate particle which, as a result of being magnetized, produces a surrounding strayfield, Hp. This strayfield has to cause a detectable perturbation in the MTJ cell's state of magnetization. In sensor designs of the prior art, that detectable perturbation is a polarity reversal of the storage states, from M1 to M2 or vice versa. Such a reversal response is advantageous with a biosensor since it generates a non-ambiguous, storable detection result. The drawback is that a comparatively large magnetized particle is required in order for its strayfield Hp>Hc. This, in turn, leads to a problem. because large particles are more difficult to manipulate in the analyte (the fluid containing the biological particles being identified). Currently, the compromise particle size is approximately 1 micron.
Because the strayfield, Hp, produced by the magnetized particle is fairly small, it is imperative to design MTJ sensors that have a high sensitivity. This is usually achieved by producing sensors with as low a magnetic anisotropy as possible, so that the magnetization is easily changed in direction, but is not too unstable to allow for storage. With such low anisotropy, however, the variations from one MTJ to another become significant and difficult to control. Therefore, it is difficult to design MTJ sensors that can reliably and consistently detect small magnetized beads.
Thus, we see there are several mutually conflicting requirements to constructing an efficient biosensor device based on MTJ cell technology or, for that matter, based on any technology (not necessarily MTJ technology) in which the sensor operates on the basis of a bi-stable state.
In MTJ technology high sensitivity requires low anisotropy. But stability and storage requires high anisotropy. High anisotropy, in turn, requires large particles for detection, since they produce large strayfields. But large particles are difficult to maneuver within an analyte. We shall see below how the present invention solves these problems.
Given the increasing interest in the identification of biological molecules, it is to be expected that there is a significant amount of prior art directed at the use of magnetic MTJ cell sensors (and other magnetic sensors) to provide this identification. An early disclosure of the use of magnetic labels (magnetized particles) to detect target molecules is to be found in Baselt (U.S. Pat. No. 5,981,297). Baselt describes a system for binding target molecules to recognition agents that are themselves covalently bound to the surface of a magnetic field sensor. The target molecules, as well as non-target molecules, are covalently bound to magnetizable particles. The magnetizable particles are preferably superparamagnetic iron-oxide impregnated polymer beads and the sensor is a magnetoresistive material. The detector can indicate the presence or absence of a target molecule while molecules that do not bind to the recognition agents (non-target molecules) are removed from the system by the application of a magnetic field.
A particularly detailed discussion of the detection scheme of the method is provided by Tondra (U.S. Pat. No. 6,875,621). Tondra teaches a ferromagnetic thin-film based GMR magnetic field sensor for detecting the presence of selected molecular species. Tondra also teaches methods for enhancing the sensitivity of magnetic sensor arrays that include the use of bridge circuits and series connections of multiple sensor stripes. Tondra teaches the use of paramagnetic beads that have very little intrinsic magnetic field and are magnetized by an external source after the target molecules have been captured.
Prinz et al. (U.S. Pat. Nos. 6,844,202 and 6,764,861) teaches the use of a magnetic sensing element in which a planar layer of electrically conducting ferromagnetic material has an initial state in which the material has a circular magnetic moment. In other respects, the sensor of Prinz fulfills the basic steps of binding at its surface with target molecules that are part of a fluid test medium. Unlike the GMR devices disclosed by Tondra above, the sensor of Prinz changes its magnetic moment from circular to radial under the influence of the fringing fields produced by the magnetized particles on the bound target molecules.
U.S. Pat. No. 7,031,186 and Patent Application 2004/0120185 (Kang et al) disclose a biosensor comprising MTJ elements.
U.S. Patent Application 2007/0159175 (Prinz) shows on-chip magnetic sensors to detect different types of magnetic particles or molecules.
U.S. Patent Application 2007/0114180 (Ramanathan et al) teaches MTJ channel detectors for magnetic nanoparticles.
U.S. Patent Application 2005/0100930 (Wang et al) discloses detection of biological cells and molecules.
None of the prior art inventions cited above provide a robust method of reliably detecting the presence of small magnetized particles bonded to biological molecules. It is the object of the present invention to provide such a method that has improved sensitivity so as to be able to reliably detect reduced size particles.